A liquid chromatography apparatus for pressurizing a sample with liquid as a mobile phase so as to send them to a column, separating and eluting components of the sample and detecting them on a detector basically has a configuration as shown in FIG. 4. In FIG. 4, the liquid chromatography apparatus includes a mobile phase reservoir 101 for storing the liquid as a mobile phase, a mobile phase degassing device 102 for removing air from the liquid as a mobile phase, a pump 103 for sending the liquid as a mobile phase from the mobile phase reservoir 101 to a detector 107, an (automatic) sample injection apparatus 104 for injecting a sample into the liquid as a mobile phase directed into a separation column 105, the separation column 105 being filled with a packing material for separating components of the sample, a column thermostatic bath 106 for keeping the temperature of the separation column 105 to be approximately constant, and the detector 107 for detecting an eluted component of the sample.
As the detection sensitivity of a liquid chromatography apparatus having such a structure becomes higher, a phenomenon called carry-over has become problematic. The carry-over is a phenomenon such that a substance of a sample measured previously through time remains in the liquid chromatography apparatus so as to show a detection result as if the substance were present in the sample measured at present, and reduces the reliability of the results of analysis. The carry-over is caused, when a sample is adsorbed on a metal and/or a resin and remains in the automatic sample injection apparatus 104 after the sample is injected into the liquid as a mobile phase in the automatic sample injection apparatus 104, and the remaining sample is introduced into the liquid chromatography analysis system when a next sample is introduced. Particularly, it is known that a sample containing a basic and/or fat-soluble substance is easily adsorbed on the metal and/or resin in the automatic sample injection apparatus 104 and, therefore, easily remains in the automatic sample injection apparatus 104 so that carry-over is significantly observed.
In order to reduce the carry-over by removing the sample adsorbed and remaining in the automatic sample injection apparatus 104, a self-cleaning function can be provided in the automatic sample injection apparatus 104. As shown in FIG. 6, the main operation of the automatic sample injection apparatus 104 is composed of sampling a sample supplied into a sampling vessel 14 by a sampling needle 10, inserting the sampling needle 10 in which the sample is sampled into an injection port 19 of an injection valve 15 so as to inject the sample from the sampling needle 10 through a sample injection route to a sample loop 16, and switching flow channels for a sample in the injection valve 15 so as to send the sample in the sample loop 16 to the separation column 105. Therefore, as locations (sample adsorbing portions) at which the sample in the automatic sample injection apparatus 104 is adsorbed and remains, an outer wall of the sampling needle 10 after sampling the sample, an inner wall of the sampling needle 10 after sampling the sample, an inner wall of the sample loop 16 after sending the sample to the separation column 105, and the flow channel for a sample in the injection valve 15 are listed.
Since the self-cleaning functions are provided to these sample adsorbing portions, it is only necessary to provide four self-cleaning functions, that is, a function for cleaning the outer wall of the sampling needle, a function for cleaning the inner wall of the sampling needle, and functions for cleaning the sample loop and the injection valve. These four self-cleaning functions are described below.
(1) The function for cleaning the outer wall of the sampling needle immediately before sample injection (Needle pre-washing)
As shown in FIG. 7, needle pre-washing is a function for removing a sample adsorbed and remaining on the outer wall of the sampling needle 10 after sampling the sample by dipping the sampling needle 10 into cleaning liquid that is sent into a cleaning part 17 immediately before the sample sampled in the sampling needle 10 is injected into an injection port. The cleaning liquid used for removing the sample remaining on the outer wall of the sampling needle 10 is disposed of and pure cleaning liquid is supplied into the cleaning part 17. For the needle pre-washing, a user can set a time period (cleaning time period) for dipping the sampling needle 10 in the cleaning liquid.
(2) The function for cleaning the inner wall of the sampling needle immediately after sample injection (Needle post-washing)
As shown in FIG. 8, needle post-washing is a function for removing a sample adsorbed and remaining on the inner wall of the sampling needle 10 by dipping the sampling needle 10 into cleaning liquid supplied into the cleaning part 17 and flowing the cleaning liquid through the sampling needle 10 immediately after the sample is injected into the injection port. The cleaning liquid used for removing the sample remaining on the inner wall of the sampling needle 10 is disposed of and pure cleaning liquid is supplied into the cleaning part 17. For the needle post-washing, a user can set a time period (cleaning time period) for flowing the cleaning liquid through the sampling needle 10.
(3) The cleaning of the sample injection route after the sample injection (Post-injection washing)
As show in FIG. 9, the post-injection washing is a cleaning function in which after a sample is injected into an injection port, operations (1) through (3) are carried out in order during the analysis of the sample.
1) The sampling needle 10 is moved to a position above a cleaning port 24 and cleaning liquid prepared in the cleaning port 24 is aspirated into the sampling needle 10.
2) The sampling needle 10 that aspirated and holds the cleaning liquid is inserted into the injection port 19 of the injection valve 15 and the cleaning liquid in the sampling needle 10 is (repeatedly) ejected and aspirated so as to clean a sample injection route (flow channel from the injection port 19 to a waste liquid disposal port 23) in the injection valve 15. At this time, the injection port 19 and the waste liquid disposal port 23 are connected and the injection port 19 is not connected to the sample loop 16 in the injection valve 15. Therefore, as the injection port 19 and the waste liquid disposal port 23 are connected after injecting a sample into the sample injection route, while the sample is analyzed through the sample loop 16, a flow channel between the injection port 19 and the waste liquid disposal port 23 can be cleaned.
3) For the post-injection washing, a user can set the volume of cleaning liquid to be aspirated into the sampling needle 10 and the number of ejections and aspirations of the cleaning liquid (the number of cleanings). Also, two or more cleaning liquid ports 24 can be provided so as to use two or more kinds of cleaning liquids. For example, when the two or more kinds of cleaning liquids are used, if a previously used cleaning liquid is liquid with high detergency (a strong alkali that is not desired to pass through the separation column 15, etc.) and a subsequently used cleaning liquid is liquid to be used as a mobile phase, the liquid with high detergency can be used and the liquid with high detergency can be prevented from mixing into an analysis system.
(4) A function of cleaning the sample loop and the injection valve during the progression of analysis (Loop rinse)
For performing the loop rinse, a loop rinse valve is required except for the automatic sample injection apparatus. As shown in FIGS. 10(a) and 10(b), a loop rinse valve 108 operates so as to switch flow channels of liquid as a mobile phase supplied from the mobile phase reservoir 101. That is, as shown in FIG. 10(a), when the cleaning function of the loop rinse is not performed, the liquid as the mobile phase sent out by the pump 103 is supplied into the sample injection apparatus 104 and sent to the separation column 105 and the detector 107 with a sample. On the other hand, as shown in FIG. 10(b), when the cleaning function of the loop rinse is performed, the liquid as the mobile phase sent out by the pump 103 does not pass through the sample injection apparatus 104 and is directly sent to the separation column 105 and the detector 107. As such a loop rinse valve 108 is used, if after sending the sample to the separation column 105 through a flow channel shown in FIG. 10(a), immediately switching to a flow channel shown in FIG. 10(b) is made, while analysis is progressed by sending the liquid as the mobile phase to the separation column 105, the sample loop and the injection valve can be cleaned.
The loop rinse is a cleaning function for carrying out the following operations 1) through 4) in order after injecting a sample into the sample loop 16, as shown in FIG. 10 and FIG. 11.
1) After the sample is injected into the sample loop 16, the automatic sample injection apparatus is completely separated from a flow channel of an analysis system including the separation column 105 and the detector 107 by the loop rinse valve 108.
2) While analysis is carried out by sending the liquid as the mobile phase to the separation column without traveling through the automatic sample injection apparatus 104, the insides of the sample loop 16 and injection valve 15 are cleaned by inserting the sampling needle 10 into the injection port 19 of the injection valve 15 and ejecting the cleaning liquid from the sampling needle 10 in the automatic sample injection apparatus 104. Then, the sampling needle 10 is connected to a cleaning liquid reservoir in which the cleaning liquid is stored and the cleaning liquid is supplied to the sampling needle from the cleaning liquid reservoir.
3) Since the flow channels of the injection valve 15 are switched while the cleaning liquid is ejected from the sampling needle, a flow channel from the injection port 19 to the waste liquid disposal port 23 can be also cleaned.
4) As the cleaning in the sample loop 16 and the injection valve 15 is completed, the loop rinse valve 108 is switched to the usual flow channel through the automatic sample injection apparatus 104, again, as shown in FIG. 10(a).
For the loop rinse, a user can set a time period of ejecting the cleaning liquid from the sampling needle 10 (cleaning time period) and the number of times of switching of the injection valve 15. Also, two or more vessels for cleaning liquid can be provided so that two or more kinds of cleaning liquids can be used by connecting these vessels for cleaning liquid to the sampling needle 10 and switching the connections among these vessels for cleaning liquids and the sampling needle 10. For example, when the two or more kinds of cleaning liquids are used, if previously used cleaning liquid is liquid with high detergency (a strong alkali that is not desired to pass through the separation column 105, etc.) and subsequently used cleaning liquid is liquid to be used as a mobile phase, the liquid with high detergency can be used and the liquid with high detergency can be prevented from mixing into an analysis system.
On the other hand, in regard to cleaning of a needle, a sample introduction apparatus for introducing a sample to an analyzer such as a liquid chromatography apparatus, which sample introduction apparatus can perform the mixing of samples or the cleaning of needle without aspirating or ejecting liquid, is also disclosed in JP-A-11-304779. This sample introduction apparatus is equipped with a vibration generation part such as an ultrasonic vibrator arranged directly or through the intermediary of a member capable of transmitting the vibration, on a sample injection needle, and a vibration control part for controlling the vibration generation part. More specifically, it is equipped with an ultrasonic vibrator arranged in connection with the sample injection needle or a metal part connecting to the sample injection needle, and the needle itself is vibrated by the ultrasonic vibrator which is simultaneously controlled by the vibration control part, at the time of mixing the samples or cleaning the needle.
Since the automatic sample injection apparatus provided with four self-cleaning functions performs the four self-cleaning functions in combination in order to reduce a carry-over phenomenon, a cleaning time period of approximately 3 minutes on average is needed. Among these four self-cleaning functions, the needle pre-washing is performed before the start of analysis but cleaning for approximately 1 through 5 seconds is commonly sufficient so that a shorter cleaning time period is required. Therefore, the three self-cleaning functions other than the needle pre-washing require approximately 3 minutes in total. However, these three self-cleaning functions are practicable while a sample is analyzed. Accordingly, if the time period of analysis is 3 minutes or more, all the three self-cleaning functions are carried out within the time period of analysis, in which there is no problem. However, in regard to analysis by means of liquid chromatography, the time period of analysis is being reduced and the cleaning time period has to be reduced (for example, to a cleaning time period=1 minute or less) in accordance with the reduction of the time period of analysis.
Also, it is necessary to set the operating conditions of the four self-cleaning functions, respectively, in the automatic sample injection apparatus provided with the four self-cleaning functions, in order to set the optimum cleaning conditions adapted to a sample to be analyzed. However, as described above, since it is necessary to set approximately 6 through 8 parameters in total for the operating conditions of the four self-cleaning functions, the number of the parameters to be set is high, which is complex and inconvenient for a user.
On the other hand, in regard to the sample introduction apparatus provided with a vibration generation part such as an ultrasonic vibrator arranged directly or through the intermediary of a member capable of transmitting the vibration, on a sample injection needle, since the sample injection needle it self is vibrated, adverse impact is applied to the sample injection needle. As the result, it is a concern that the deterioration of the sample injection needle is caused and the durability of the sample injection needle is lowered. Also, as the sample injection needle is cleaned on the condition that a sample is held in such a sample injection needle, since the sample injection needle itself vibrates, a part of the sample held in the sample injection needle is lost in the cleaning liquid and there is a possibility of causing a large error in the amount of injected sample. Further, there is also a possibility of transmitting vibration through the sample injection needle to another member of the automatic sample injection apparatus by vibrating the sample injection needle itself. However, since the automatic sample injection apparatus is fundamentally a precision machine, it is required to avoid undesired vibration. Therefore, there is also a problem in that the sample introduction apparatus provided with a vibration generation part such as an ultrasonic vibrator arranged directly or through the intermediary of a member capable of transmitting the vibration on a sample injection needle, can be influenced by the ultrasonic vibration of the sample injection needle.